EP1623715A1 - Composition comportant un agent pharmacologiquement actif couplé à un élément cible spécifique de cellules et une saponine - Google Patents

Composition comportant un agent pharmacologiquement actif couplé à un élément cible spécifique de cellules et une saponine Download PDF

Info

Publication number
EP1623715A1
EP1623715A1 EP04016497A EP04016497A EP1623715A1 EP 1623715 A1 EP1623715 A1 EP 1623715A1 EP 04016497 A EP04016497 A EP 04016497A EP 04016497 A EP04016497 A EP 04016497A EP 1623715 A1 EP1623715 A1 EP 1623715A1
Authority
EP
European Patent Office
Prior art keywords
active agent
target cell
pharmacologically active
saponin
specific component
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Granted
Application number
EP04016497A
Other languages
German (de)
English (en)
Other versions
EP1623715B1 (fr
Inventor
Hendrik Fuchs
Mark Sutherland
Christopher Bachran
Matthias Melzig
Iring Heisler
Philip Hebestreit
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Charite Universitaetsmedizin Berlin
Freie Universitaet Berlin
Original Assignee
Charite Universitaetsmedizin Berlin
Freie Universitaet Berlin
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Charite Universitaetsmedizin Berlin, Freie Universitaet Berlin filed Critical Charite Universitaetsmedizin Berlin
Priority to EP04016497A priority Critical patent/EP1623715B1/fr
Priority to ES04016497T priority patent/ES2395818T3/es
Priority to JP2007520738A priority patent/JP2008505958A/ja
Priority to US11/630,752 priority patent/US20080064762A1/en
Priority to CA002571739A priority patent/CA2571739A1/fr
Priority to PCT/EP2005/007557 priority patent/WO2006005581A2/fr
Publication of EP1623715A1 publication Critical patent/EP1623715A1/fr
Application granted granted Critical
Publication of EP1623715B1 publication Critical patent/EP1623715B1/fr
Anticipated expiration legal-status Critical
Expired - Lifetime legal-status Critical Current

Links

Images

Classifications

    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
    • A61K36/18—Magnoliophyta (angiosperms)
    • A61K36/185—Magnoliopsida (dicotyledons)
    • A61K36/36—Caryophyllaceae (Pink family), e.g. babysbreath or soapwort
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
    • A61K47/51—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent
    • A61K47/62—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being a protein, peptide or polyamino acid
    • A61K47/64—Drug-peptide, drug-protein or drug-polyamino acid conjugates, i.e. the modifying agent being a peptide, protein or polyamino acid which is covalently bonded or complexed to a therapeutically active agent
    • A61K47/6415—Toxins or lectins, e.g. clostridial toxins or Pseudomonas exotoxins
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
    • A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
    • A61K47/51—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent
    • A61K47/62—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being a protein, peptide or polyamino acid
    • A61K47/64—Drug-peptide, drug-protein or drug-polyamino acid conjugates, i.e. the modifying agent being a peptide, protein or polyamino acid which is covalently bonded or complexed to a therapeutically active agent
    • A61K47/6425—Drug-peptide, drug-protein or drug-polyamino acid conjugates, i.e. the modifying agent being a peptide, protein or polyamino acid which is covalently bonded or complexed to a therapeutically active agent the peptide or protein in the drug conjugate being a receptor, e.g. CD4, a cell surface antigen, i.e. not a peptide ligand targeting the antigen, or a cell surface determinant, i.e. a part of the surface of a cell
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P25/00—Drugs for disorders of the nervous system
    • A61P25/28—Drugs for disorders of the nervous system for treating neurodegenerative disorders of the central nervous system, e.g. nootropic agents, cognition enhancers, drugs for treating Alzheimer's disease or other forms of dementia
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P29/00—Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P3/00—Drugs for disorders of the metabolism
    • A61P3/08—Drugs for disorders of the metabolism for glucose homeostasis
    • A61P3/10—Drugs for disorders of the metabolism for glucose homeostasis for hyperglycaemia, e.g. antidiabetics
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
    • A61P31/04—Antibacterial agents
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
    • A61P31/12—Antivirals
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
    • A61P31/12—Antivirals
    • A61P31/14—Antivirals for RNA viruses
    • A61P31/18—Antivirals for RNA viruses for HIV
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P43/00—Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P5/00—Drugs for disorders of the endocrine system
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P9/00—Drugs for disorders of the cardiovascular system

Definitions

  • the present invention relates to a composition comprising a pharmacologically active agent coupled to a target cell specific component, and a saponin, to uses of said composition and to a kit comprising said composition.
  • the invention also relates to a method of enhancing the function and/or the target cell specificity of a pharmacologically active agent. It furthermore relates to uses of a saponin.
  • immunotoxins are chimeric proteins or chemically coupled conjugates in which a cell-targeting moiety is combined with a cytotoxic agent.
  • a cell-targeting moiety is either a small radioactive molecule, a low molecular weight organic compound or the catalytic domain of a natural toxin.
  • the cell targeting moiety is an antibody fragment or a natural ligand for cancer-associated endocytic surface antigens.
  • Typical target antigens are the epidermal growth factor (EGF) receptor, the proto-oncogene receptor ErbB2 (also known as HER-2 in humans), the interleukin-2 receptor or cancer-associated carbohydrates (1).
  • the first recombinant IT approved by the American Food and Drug Administration was Ontak. It consists of interleukin-2 and parts of diphtheria toxin. It was developed for the treatment of cutaneous T-cell lymphoma (2, 3). In addition to Ontak two further chemically coupled ITs, Zevalin (4, 5) and Mylotarg (6), have been approved and several ITs are under investigation in clinical trials.
  • the natural membrane translocating sequence of the toxin is used to mediate endosomal membrane transfer. While this is possible e.g. for diphtheria toxin and Pseudomonas exotoxin other toxins like plant type I ribosome-inactivating proteins (RIP) lacking a translocating sequence must enter the cells using other to date unknown mechanisms.
  • PTD protein transduction domains
  • Trojan peptides or cell permeable proteins capable of transporting effector molecules into cells has become an increasingly attractive mechanism to solve this problem (reviewed in (10, 11)).
  • ITs such as BL22 or Ontak rely on the translocation domain of the natural toxin for effective uptake (14). This limits the number of natural toxins available for the construction of ITs.
  • toxins that do not possess a translocation domain like diphtheria toxin A-chain, ricin toxin A-chain, the type 1 RIP saporin-3 or toxic human proteins the introduction of membrane permeable peptides, called protein transduction domains (PTD) or Trojan peptides, was suggested (8).
  • PTD protein transduction domains
  • a composition in which the specific function of a pharmacologically active agent, in particular the cytotoxicity of a chimeric toxin or a toxin conjugate, on target cells is increased. Furthermore it was an object of the present invention to reduce the side-effects associated with the use of pharmacologically active agents, for example during tumor therapy. Furthermore it was an object of the present invention to provide for ways by which the amount of pharmacologically active agents to be used and administered in a therapy can be substantially reduced.
  • compositions comprising at least one pharmacologically active agent coupled to a target cell specific component, and at least one saponin, wherein said pharmacologically active agent is not a saponin, and is not an antigen binding protein comprising an antigen binding region and a region or regions of an antibody that mediate antibody dependent immunological processes.
  • said pharmacologically active agent is not an agent stimulating an immune response.
  • said pharmacologically active agent is able to eliminate a dysfunction of a cell or to kill a cell by causing target cell death, preventing target cell proliferation or abolishing or reducing the dysfunction of the target cell by interaction with intracellular targets.
  • said pharmacological active agent is cytotoxic or cytostatic.
  • said target cell specific component binds specifically to a target cell and is selected from the group comprising (a) proteins, such as natural proteins, recombinant proteins, antibodies, antibody fragments, lipoproteins and protein ligands, such as growth factors (e.g. epidermal growth factor (EGF), granulocyte-macrophage colony-stimulating factor (GM-CSF), vascular endothelial growth factor (VEGF), transforming growth factor (TGF)) and chimeras thereof, cytokines (e.g. interleukins, interferones), transferrin, adhesion molecules (e.g.
  • proteins such as natural proteins, recombinant proteins, antibodies, antibody fragments, lipoproteins and protein ligands, such as growth factors (e.g. epidermal growth factor (EGF), granulocyte-macrophage colony-stimulating factor (GM-CSF), vascular endothelial growth factor (VEGF), transforming growth factor (TGF)) and
  • CD4 urokinase-type plasminogen activator
  • peptides such as natural peptides and synthetic peptides
  • nucleic acids such as natural or designed DNA, RNA or derivatives thereof
  • carbohydrates such as natural or designed DNA, RNA or derivatives thereof
  • lipids such as natural or designed lipids
  • hormones and chemical compounds including such designed by combinatorial chemistry from libraries and combinations of (a) - (g).
  • target cell is selected from the group comprising cancerous cells, parasitic cells, cells infected by a virus, and cells releasing TNF ⁇ .
  • said pharmacologically active agent is coupled to said target cell specific component via non-covalent association and/or via covalent linkage.
  • said target cell specific component is an antibody or an antibody fragment or a growth factor, cytokine or transferrin, or a combination or derivative of any of these.
  • said target cell specific component binds to a receptor selected form the group comprising growth factor receptors (e.g. HER-1 (epidermal growth factor receptor EGFR), HER-2 HER-3, HER-4 or granulocyte-macrophage colony-stimulating factor receptor GM-CSFR), cytokine receptors (e.g. IL-2R, IL-25R), transferrin receptors, cluster of differentiation (CD) antigens (e.g. CD5, CD6, CD7, CD19, CD22, CD25, CD30, CD33, CD56), Lewis antigens (e.g. Le y ) or Wilms' tumor gene product.
  • growth factor receptors e.g. HER-1 (epidermal growth factor receptor EGFR), HER-2 HER-3, HER-4 or granulocyte-macrophage colony-stimulating factor receptor GM-CSFR
  • cytokine receptors e.g. IL-2R, IL-25R
  • said pharmacologically active agent is a cytotoxin.
  • said cytotoxin is selected from the group comprising (a) toxic plant proteins, such as ribosome-inactivating proteins (RIP), in particular type I proteins (e.g. saporin, dianthin, gelonin, PAP) and enzymatical active fragments thereof and the A chain of type II proteins (e.g.
  • RIP ribosome-inactivating proteins
  • type I proteins e.g. saporin, dianthin, gelonin, PAP
  • enzymatical active fragments thereof e.g.
  • ricin from ricin, abrin, mistletoe lectin (viscumin), modeccin) but not excluding other fragments or full length type II proteins
  • toxic fungal proteins and peptides such as ⁇ -sarcin or mitogillin
  • bacterial toxins such as diphtheria toxin or Pseudomonas exotoxin
  • toxic animal proteins such as angiogenin or granzyme B, in particular of human origin
  • radioactive compounds and combinations or derivatives of (a) - (e).
  • said pharmacologically active agent coupled to said target cell specific component is a chimeric toxin or toxin conjugate, wherein, preferably, said pharmacologically active agent is a peptidic or protein cytotoxin and said target cell specific component is an antibody, growth factor, cytokine or transferrin or a fragment, combination or derivative thereof.
  • said pharmacologically active agent coupled to said target cell specific component is a single recombinant protein.
  • said saponin is selected from the group comprising steroid saponins and triterpenoic saponins, preferably having an aldehyde function at the aglycone and two sugar residues glycosidically bound to the aglycone.
  • said saponin is selected from the group comprising triterpenoic saponins with basic structures belonging to the type of 12,13-dehydrooleanane with an aldehyde function in position 23, e.g. Saponinum album from Gypsophila paniculata.
  • said chimeric toxin or toxin conjugate is selected from the group comprising OntakTM (DAB 389 IL2), Zevalin (ibritumomab tituxetan), Mylotarg (gemtuzumab ozogamicin), Saporin-EGF, Herceptin-DM1 (trastuzumab-DM1), RFB4-PE38 (BL22), DAB 389 EGF, H65-RTA, Anti-CD6-bR, Anti-CD7-dgA, TXU-PAP, Anti-Tac-PE38, RFT5-SMPT-dgA, IgG-RFB4-dgA, Di-dgA-RFB4, IgG-HD37-dgA, Anti-B4-bR, Anti-My9-bR, DT388-GM-CSF, B3-LysPE38, B3-PE38, TP40, 454A12-rRA, Tf-CRM107
  • chimeric toxins are described in references (9, 16-19) or are commcercially available from or sponsored by Seragen (Ontak), IDEC Pharmaceuticals (Zevalin), Wyeth Laboratories (Mylotarg), Genentech (Herceptin-DM1), US National Cancer Institute (BL22), Corixa, GlaxoSmith-Kline (Bexxar), Antisoma (Theragyn).
  • said composition further comprises at least one of the following functional units: a transport protein capable of transporting said pharmacologically active agent into a cell, a cytosolic cleavable bond, preferably a cytosolic cleavable peptide, an endosomal cleavable bond, preferably an endosomal cleavable peptide wherein, preferably, said pharmacologically active agent is coupled to said target cell specific component via said transport protein, and wherein, more preferably, said transport protein is a protein transduction domain (PTD).
  • PTD protein transduction domain
  • a pharmacologically active protein coupled to a target cell specific component via at least one of the following functional units: a protein transduction domain or a cytosolic cleavable peptide or an endosomal cleavable peptide, is herein also sometimes referred to as an "immunoadapter agent", or more specifically “immunoadapter protein”.
  • composition according to the present invention for use as a medicament.
  • composition according to the present invention for the manufacture of a medicament for the treatment of an animal having a cancerous, infectious, cardiovascular or inflammatory disease or diabetes.
  • said saponin is administered separately from said pharmacologically active agent coupled to said target cell specific component.
  • said saponin is administered via a different route to the administration route of said pharmacologically active agent coupled to said target cell specific component, wherein, preferably, said saponin is administered via injection and said pharmacologically active agent coupled to said target cell specific component is administered via ingestion or vice versa.
  • said saponin is administered via the same route as said pharmacologically active agent coupled to said target cell specific component, wherein, preferably, said administration occurs via injection.
  • said saponin is administered before said pharmacologically active agent coupled to said target cell specific component, wherein, preferably, said saponin is administered one minute to one hour, preferably one minute to 45 minutes, more preferably one minute to 30 minutes, most preferably 5 minutes - 20 minutes before administration of said pharmacologically active agent coupled to said target specific component.
  • said saponin is administered at an amount of 100 ⁇ g - 100 mg per kg bodyweight of said animal.
  • said animal is a human being.
  • kits comprising, in one or more containers, said composition according to the present invention.
  • said pharmacologically active agent coupled to said target cell specific component, and said saponin are in separate containers.
  • said pharmacologically active agent coupled to said target cell specific component is formulated for injection and said saponin is also formulated for injection.
  • the objects of the present invention are also solved by a method of enhancing the function and/or the target cell specificity of a pharmacologically active agent which pharmacologically active agent is not a saponin, and is not an antigen binding protein comprising an antigen binding region and a region that mediates antibody dependent immunological processes, said method comprising coupling said pharmacologically active agent to a target cell specific component, and combining said pharmacologically active agent coupled to said target cell specific component with at least one saponin.
  • said pharmacologically active agent is able to eliminate a dysfunction of a cell or to kill a cell including parasites, in an animal having a disease caused by said dysfunction or parasite, by causing target cell death, preventing target cell proliferation or abolishing or reducing the dysfunction of the target cell by interaction of said pharmacological active agent with intracellular targets involved directly or indirectly in said dysfunction, wherein said dysfunction comprises, but is not limited to, such caused by infection (e.g. viral infection such as a HIV or bacterial infection) or such leading to or being associated with cancer, cardiovascular diseases, inflammation, neurodegenerative and/or amyloid diseases, or diabetes.
  • infection e.g. viral infection such as a HIV or bacterial infection
  • said dysfunction comprises, but is not limited to, such caused by infection (e.g. viral infection such as a HIV or bacterial infection) or such leading to or being associated with cancer, cardiovascular diseases, inflammation, neurodegenerative and/or amyloid diseases, or diabetes.
  • said pharmacologically active agent is not an agent stimulating an immune response.
  • said function is a cytotoxic or cytostatic function.
  • said disease is selected from the group comprising cancerous diseases, infectious diseases, inflammatory diseases, diseases of the cardiovascular system, and diabetes.
  • said combining occurs by packaging said pharmacologically active agent coupled to said target cell specific component and said saponin together, albeit, optionally, in separate containers.
  • said pharmacologically active agent, said target cell specific component and said saponin are as defined above.
  • the objects of the invention are also solved by the use of a saponin as defined above for enhancing the function and/or the target cell specificity of a pharmacologically active agent, as defined above, said pharmacologically active agent being coupled to a target cell specific component, as defined above, for the manufacture of a medicament for treatment of a disease selected from the group comprising cancerous diseases, infections diseases, inflammatory diseases, diseases of the cardiovascular system and diabetes.
  • a pharmacologically active agent which is cytotoxic, coupled to a target cell specific agent, both as defined before, in combination with a saponin, as defined before.
  • the administration may occur in any of the ways and order as described before.
  • said saponin is administered before said pharmacologically active agent.
  • the term "pharmacologically active agent” is meant to designate any agent that has a detectable pharmacological effect on a cell or an organism.
  • a pharmacologically active agent examples include toxins, cytostatica, radioactive compounds, enzymes, enzyme inhibitors, receptor ligands, hormones, transcription factors, intracellular signal molecules, antisense or small interfering nucleic acids.
  • said pharmacologically active agent are also antibodies or antibody fragments provided that their function is the elimination or reduction of said dysfunction and not to mediate an antibody-dependent immunological process.
  • the pharmacologically active agent according to the present invention is a cytotoxin or at least a cytostatic agent.
  • the pharmacologically active agent is not a saponin, and is not an antigen binding protein comprising an antigen binding region and a region or regions of an antibody that mediate antibody dependent immunological processes. In one embodiment, it is not an agent stimulating an immune response.
  • target cell specific component is any component which binds to a target cell with a higher probability, possibly because of a molecular structure on said target cell that is recognized by said target cell specific component.
  • said target cell is a cancerous cell.
  • the coupling of said pharmacologically active agent to said target cell specific component may be in the form of a non-covalently associated conjugate, a covalent linkage between the two parts or, in the case of the two parts being proteins, a recombinantly produced fusion protein.
  • said pharmacologically active agent coupled to said target cell specific component is a chimeric toxin or a toxin conjugate.
  • chimeric toxin or a toxin conjugate is meant to signify a cytotoxin coupled to an antibody, an antibody fragment or to another component, preferably a ligand, more preferably a protein ligand, such as but not limited to growth factors, cytokines or transferrin, which recognizes specific structures on a target cell such as but not limited to growth factor receptors, cytokine receptors, transferrin receptors, cluster of differentiation (CD) antigens, Lewis antigens or Wilms' tumor gene product.
  • CD cluster of differentiation
  • transport protein is meant to designate any protein or peptide that is capable of transporting a pharmacologically active agent into a cell. Examples hereof, such as PTD, have been described above.
  • immunoadaptertoxin is meant to signify a recombinant chimeric toxin containing in addition to said pharmacologically active agent and said target cell specific component a protein or peptide that contains at least one of the following functional units, a cytosolic cleavable peptide, an endosomal cleavable peptide or a transport protein, such as PTD.
  • antibody dependent immunological processes as used herein is meant to designate processes like antibody dependent cellular cytotoxicity, activation of complement, opsonization and phagocytosis.
  • dysfunction of a cell is meant to designate any deviation of the cell from its normal healthy state. Such dysfunction of a cell may for example manifest itself in a loss of function, loss of differentiation, increased or decreased metabolic turnover, loss of capability of undergoing apoptosis, premature apoptosis, and others.
  • parasite is meant to designate any agent or organism which makes use of a host cell's metabolism, lives off a host cell's substance of matter and/or interferes with a host cell and its functions in a manner deleterious to a host cell.
  • the term includes, but is not limited to viruses, bacteria and protozoa.
  • saponins e.g. gypsophilasaponin
  • Saponinum album from Gypsophila paniculata was used as an example, it is bisdesmosidic and has Gypsogenin as aglycone.
  • saponins are glycosidic compounds which have one non-sugar component, also termed "aglycone", and one to several sugar residues attached to said aglycone.
  • Saponins having one sugar residue are called monodesmosidic, saponins having two sugar residues attached are termed bisdesmosidic, three sugar residues tridesmosidic etc..
  • Saponins can be classified according to the type of aglycone they have.
  • the two main types of saponins are steroid saponins and triterpenoic saponins.
  • Many of the triterpenoic saponins are of the oleanane type which is a triterpenoic ring structure having 30 carbon atoms.
  • both ligand-free Sap-3 on target cells as well as complete ITs on receptor-free cells showed a drastically lower cytotoxicity revealing that the effect is receptor- and ligand-specific.
  • the Spn-mediated enhancement of the unspecific toxicity is solely 490 to 1450-fold and independent of the cell line.
  • the high target specificity of saponin-enhanced ITs permits an application of the IT in a broad range and therefore an adjustment to the number of receptors on the target cell.
  • Quillaja saponins are a permitted food additive and long-term feeding to rats (0.75 % of total diet) showed no effects on food consumption or body weight (21).
  • Ryokucha saponin one of the ingredients of green tea (Camellia sinensis), had no effect on rats fed for 30 days with 50 mg per kg per day (22). These saponins even show an anti-allergic effect on rats and guinea pigs (23).
  • saponins have not only been tested orally but also intravenously. Consecutive intravenous administrations of ginseng saponins, up to 500 ⁇ g per mouse per day, revealed an inhibitory effect on tumor angiogenesis and metastasis but did not generate severe side effects (24, 25).
  • a dose of approximately 30 ⁇ g to a 20 g mouse (weight per weight) corresponding to an optimal saponin concentration (1.5 ⁇ g/ml) is sufficient and far below the dose of 500 ⁇ g administered by Sato et al . (25).
  • the present invention demonstrates that, surprisingly, the combined administration of individually non-toxic concentrations of an IT and a saponin leads to a highly cytotoxic effect specific for targeted tumor cells and for the first time greater than the cytotoxicity of natural diphtheria toxin for its target cells.
  • the effect is more than additive which was unexpected.
  • saponins are combined with immunoadaptertoxins instead of conventional ITs further reduction of side effects are likely in vivo due to intracellular removal of the ligand and the PTD from the toxin. After cell death the potential of the released toxin entering non-target cells should be drastically reduced since the toxin does not bear a ligand and the enhancer effect of saponins is time-limited (Fig. 3).
  • the extremely reduced effective dose can prevent vascular leak syndrome, a dose-dependent side effect of IT therapy characterized by an increase in vascular permeability resulting in interstitial edema and organ failure (7).
  • Other critical side effects resulting from high systemic concentrations and long-term use like severe immune response may also be avoided.
  • EGF epidermal growth factor
  • EGFR EGF receptor
  • IT immunotoxin
  • PBS phosphate-buffered saline
  • PTD protein transduction domain
  • RIP ribosome-inactivating protein
  • Sap-3 His-tagged saporin-3
  • Spn Saponinum album from Gypsophila paniculata L
  • SE direct fusion protein of Sap-3 and EGF
  • SA2E EGF linked to Sap-3 via a cleavable molecular adapter.
  • the adapter is composed of a cytosolic cleavable peptide (YVHDEVDRGP) containing caspase cleavage sites and a yeast recognition sequence for cytosolic cleavage, a PTD (PLSSIFSRIGDP) derived from the PreS2-domain of hepatitis B virus surface antigen (26) and an endosomal cleavable peptide (RHRQPRGNRVGRS) containing the Pseudomonas exotoxin and a modified diphtheria toxin cleavage site.
  • YVHDEVDRGP cytosolic cleavable peptide
  • PTD PLSSIFSRIGDP
  • RHRQPRGNRVGRS endosomal cleavable peptide
  • the pellets were lysed under denaturing conditions and Sap-3 as well as SA2E purified by nickel-nitrilotriacetic acid agarose chromatography according to the manufacturers' recommendations (Qiagen, Hilden, Germany). Briefly, the cells were sonicated on ice three times for 30 s (Branson sonoplus, microtip SH213G, duty cycle 20%) with 1 min breaks and centrifuged at 48000 g for 30 min at 4°C. The supernatant was subjected to affinity chromatography on a 4 ml pre-equilibrated nickel-nitrilotriacetic acid column.
  • the toxins were eluted using 8 M urea (pH 4.5) and the corresponding fractions dialyzed against 50 mM Tris-HCl / 100 mM NaCl (pH 7.5). The final materials were estimated to be 95% pure as evaluated by Coomassie and silver staining after SDS-PAGE.
  • the enzymatic activity of saporin-3 in all constructs was quantified using a colorimetric assay as described (27). All toxins used in the present study exhibited in vitro an identical N-glycosidase activity on nucleic acid substrates. The concentration of purified protein was estimated using Advanced Protein Assay Kit (Cytoskeleton Inc.). Quantity of full-length toxins was corrected after analyses by SDS-PAGE [12% (w/v) gel] under reducing conditions and densitometric scanning using protein standards.
  • HER14 cells a kind gift from Prof. E. J. van Zoelen, Department of Cell Biology, University of Nijmegen, The Netherlands
  • MCF-7 human breast adeno-carcinoma cell line
  • Cells were maintained in Dulbecco's MEM with GlutamaxTM 1 (Invitrogen /Gibco, Düsseldorf, Germany) supplemented with 10% FCS (BioChrom KG, Berlin, Germany), 100 U/ml penicillin and 100 ⁇ g/ml streptomycin. Cells were cultivated at 37°C, 5% CO 2 and 95% humidity.
  • the cells were trypsinated, washed 3x with PBS, seeded in 96-well plates (pretreated with 0.1% gelatin in PBS) at a concentration of 5000 cells per well and left untreated for 16 h.
  • the cells were washed twice with PBS and incubated with 180 ⁇ l medium containing 1.5 ⁇ g/ml Saponinum album from Gypsophila paniculata L. (Spn) except when varied as indicated in the figures.
  • Spn was purchased from Merck AG, Germany. After 5 min 20 ⁇ l toxin (Sap-3 or SA2E) was added to defined final concentrations generally in the range of 0.0001 to 30 nM and the cells cultivated for 48 h.
  • Spn was preincubated for 5 min or 1 h and then either washed out by tandem wash with PBS or left on the cells together with the toxins.
  • the toxins were preincubated for 5 min or 1 h and then either washed out or incubated together with Spn (added from a stock solution of 3 mg/ml).
  • Sap-3 and Spn were premixed and added to 180 ⁇ l medium.
  • the cytotoxicity assay is based on the cleavage of fluorescein diacetate by living cells and was performed as described by Nygren et al . (28). After incubation of the cells with the toxins, they were washed twice with PBS and incubated with fluorescein diacetate (10 ⁇ g/ml; Sigma, Germany) for 1 h. Fluorescence was measured using a microplate reader (Spectra Max Gemini, Molecular Devices) with an excitation and emission ⁇ of 485 nm and 538 nm respectively. The survival index was calculated after blank subtraction (wells without cells) as the percentage of living cells in treated wells in relation to untreated cells (cells without toxin).
  • the effect of a further increase in the Spn concentration to the toxic level of 5 ⁇ g/ml only increases the cytotoxicity to an IC 50 of 0.44 pM indicating that the extreme synergistic effect is mediated at non-toxic concentrations of Spn whereas the additional effect of toxic Spn concentrations is solely additive.
  • the same relative enhancement of the toxic effect mediated by Spn at 5 ⁇ g/ml compared to 1.5 ⁇ g/ml is also observed (at absolute higher concentrations) when applying ligand-free Sap-3 indicating that this effect is ligand-independent and therefore undesirable.
  • the present inventors used 1.5 ⁇ g/ml Spn as standard for all further investigations. Nevertheless, the present inventors also performed most experiments with 5 ⁇ g/ml Spn, the effect, however, does not differ from the effect shown in Fig. 2B and is therefore not shown for succeeding data (except for explicit Spn variations).
  • the present inventors calculated the quotient of the relative cytotoxicities determined in the presence and absence of Spn. Whereas a quotient near to one at low SA2E concentrations means that under both conditions (with and without Spn) no toxic activity can be observed, the decline of the quotient at higher concentrations is attributed to increasing cytotoxicity of SA2E in the absence of Spn. At the peak concentration the enhancer effect of Spn reaches its maximum.
  • Fig. 4B reveals that Spn shows clear effects on SA2E cytotoxicity at IT concentrations greater than 0.3 pM with a maximum at 100 pM. In contrast an impact on Sap-3 cytotoxicity is only observed at concentrations greater than 100 pM. Thus a very broad therapeutic window exists ranging from 1 to 100 pM of the IT.
  • the present inventors analyzed the Spn-mediated effect on the cytotoxicity of SA2E and Sap-3 additionally in MCF-7 cells and untransfected NIH-3T3 cells.
  • the human breast adeno-carcinoma cell line MCF-7 expresses about 40-fold less EGFR molecules than HER14 (1 ⁇ 10 4 receptors per cell compared to 4 ⁇ 10 5 ).
  • SA2E only exhibits a slight cytotoxic effect on MCF-7 cells not reaching the IC 50 at 300 nM (Fig. 6A). This corresponds to previous results and is probably due to the lower receptor number.
  • a target cell specific component e.g. a ligand
  • the concept of combined application of a saponin and a pharmacologically active agent coupled to a target cell specific component is a promising tool for enhancing the function of the pharmacologically active agent, as exemplified by the improved specific cytotoxicity of ITs on target cells and reduced side-effects during tumor therapy.

Landscapes

  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Public Health (AREA)
  • Veterinary Medicine (AREA)
  • Chemical & Material Sciences (AREA)
  • General Health & Medical Sciences (AREA)
  • Animal Behavior & Ethology (AREA)
  • Medicinal Chemistry (AREA)
  • Pharmacology & Pharmacy (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Organic Chemistry (AREA)
  • Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
  • General Chemical & Material Sciences (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Natural Medicines & Medicinal Plants (AREA)
  • Epidemiology (AREA)
  • Molecular Biology (AREA)
  • Oncology (AREA)
  • Communicable Diseases (AREA)
  • Diabetes (AREA)
  • Virology (AREA)
  • Microbiology (AREA)
  • Neurology (AREA)
  • Medical Informatics (AREA)
  • Biotechnology (AREA)
  • Mycology (AREA)
  • Alternative & Traditional Medicine (AREA)
  • Toxicology (AREA)
  • Immunology (AREA)
  • Botany (AREA)
  • Cell Biology (AREA)
  • Endocrinology (AREA)
  • Neurosurgery (AREA)
  • Biomedical Technology (AREA)
  • AIDS & HIV (AREA)
  • Obesity (AREA)
  • Heart & Thoracic Surgery (AREA)
  • Psychiatry (AREA)
EP04016497A 2004-07-13 2004-07-13 Composition comportant un agent pharmacologiquement actif couplé à un élément de ciblage spécifique de cellules et une saponine Expired - Lifetime EP1623715B1 (fr)

Priority Applications (6)

Application Number Priority Date Filing Date Title
EP04016497A EP1623715B1 (fr) 2004-07-13 2004-07-13 Composition comportant un agent pharmacologiquement actif couplé à un élément de ciblage spécifique de cellules et une saponine
ES04016497T ES2395818T3 (es) 2004-07-13 2004-07-13 Composición que comprende un agente farmacológicamente activo acoplado a un componente específico de célula diana, y una saponina
JP2007520738A JP2008505958A (ja) 2004-07-13 2005-07-12 標的細胞特異的成分に結合された薬理活性剤とサポニンとを含む組成物
US11/630,752 US20080064762A1 (en) 2004-07-13 2005-07-12 Composition Comprising a Pharmacologically Active Agent Coupled to a Target Cell Specific Component, and a Saponin
CA002571739A CA2571739A1 (fr) 2004-07-13 2005-07-12 Composition contenant un agent pharmacologiquement actif couple a un composant specifique de cellules cibles, et saponine
PCT/EP2005/007557 WO2006005581A2 (fr) 2004-07-13 2005-07-12 Composition contenant un agent pharmacologiquement actif couple a un composant specifique de cellules cibles, et saponine

Applications Claiming Priority (1)

Application Number Priority Date Filing Date Title
EP04016497A EP1623715B1 (fr) 2004-07-13 2004-07-13 Composition comportant un agent pharmacologiquement actif couplé à un élément de ciblage spécifique de cellules et une saponine

Publications (2)

Publication Number Publication Date
EP1623715A1 true EP1623715A1 (fr) 2006-02-08
EP1623715B1 EP1623715B1 (fr) 2012-09-19

Family

ID=34925740

Family Applications (1)

Application Number Title Priority Date Filing Date
EP04016497A Expired - Lifetime EP1623715B1 (fr) 2004-07-13 2004-07-13 Composition comportant un agent pharmacologiquement actif couplé à un élément de ciblage spécifique de cellules et une saponine

Country Status (6)

Country Link
US (1) US20080064762A1 (fr)
EP (1) EP1623715B1 (fr)
JP (1) JP2008505958A (fr)
CA (1) CA2571739A1 (fr)
ES (1) ES2395818T3 (fr)
WO (1) WO2006005581A2 (fr)

Cited By (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2007132481A2 (fr) * 2006-05-12 2007-11-22 Bharat Biotech International Limited Nouvelles molécules thrombolytiques et procédé associé
WO2021014026A2 (fr) 2019-07-25 2021-01-28 Sapreme Technologies B.V. Saponine bioactive liée à une fraction fonctionnelle

Families Citing this family (23)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US7524824B2 (en) * 2003-09-04 2009-04-28 Pacific Arrow Limited Composition comprising Xanthoceras sorbifolia extracts, compounds isolated from same, methods for preparing same and uses thereof
US7727561B2 (en) * 2001-08-31 2010-06-01 Pacific Arrow Limited Composition comprising Xanthoceras sorbifolia extracts, compounds isolated from same, methods for preparing same and uses thereof
US7514412B2 (en) * 2003-10-09 2009-04-07 Pacific Arrow Limited Anticancer biangeloyl saponins
US7488753B2 (en) * 2003-10-09 2009-02-10 Pacific Arrow Limited Composition comprising triterpene saponins and compounds with angeloyl functional group, methods for preparing same and uses thereof
US8614197B2 (en) * 2003-10-09 2013-12-24 Pacific Arrow Limited Anti-tumor compounds with angeloyl groups
NZ546138A (en) * 2003-10-09 2009-12-24 Pacific Arrow Ltd Composition comprising xanthoceras sorbifolia extracts, compounds isolated from same, methods for preparing same and uses thereof
US8735558B2 (en) * 2005-02-14 2014-05-27 Pacific Arrow Limited Blocking the migration or metastasis of cancer cells by affecting adhesion proteins and the uses of new compounds thereof
US9382285B2 (en) 2004-09-07 2016-07-05 Pacific Arrow Limited Methods and compounds for modulating the secretion or expression of adhesion proteins or angiopoietins of cells
US8586719B2 (en) 2005-04-27 2013-11-19 Pacific Arrow Limited Triterpenes for modulating gene expression and cell membrane, and as antiprotozoal agents
AU2007281037B2 (en) * 2006-08-03 2012-11-15 Oncology Research International Limited Methods and compositions for promoting activity of anti-cancer therapies
US9499577B2 (en) 2009-07-16 2016-11-22 Pacific Arrow Limited Natural and synthetic compounds for treating cancer and other diseases
US20120277308A1 (en) 2010-07-16 2012-11-01 Pacific Arrow Limited compounds for treating cancer and other diseases
US9434677B2 (en) 2009-07-16 2016-09-06 Pacific Arrow Limited Natural and synthetic compounds for treating cancer and other diseases
US11129906B1 (en) 2016-12-07 2021-09-28 David Gordon Bermudes Chimeric protein toxins for expression by therapeutic bacteria
US10757485B2 (en) 2017-08-25 2020-08-25 Honda Motor Co., Ltd. System and method for synchronized vehicle sensor data acquisition processing using vehicular communication
US11163317B2 (en) 2018-07-31 2021-11-02 Honda Motor Co., Ltd. System and method for shared autonomy through cooperative sensing
US11181929B2 (en) 2018-07-31 2021-11-23 Honda Motor Co., Ltd. System and method for shared autonomy through cooperative sensing
DK3773737T3 (da) * 2018-12-21 2021-10-04 Sapreme Tech Bv Saponin konjugeret til epitop-bindende proteiner
JP7281986B2 (ja) * 2019-07-05 2023-05-26 月島食品工業株式会社 飲食品の評価方法
WO2021259507A1 (fr) 2020-06-24 2021-12-30 Sapreme Technologies B.V. Conjugué d'un anticorps à domaine unique, d'une saponine et d'une molécule effectrice, composition pharmaceutique le comprenant, et utilisation thérapeutique de ladite composition pharmaceutique
CA3182793A1 (fr) 2020-06-24 2021-12-30 Ruben POSTEL Conjugue d'un anticorps a domaine unique, d'une saponine et d'une molecule effectrice, composition pharmaceutique le comprenant, et utilisation therapeutique de ladite composition pharmaceutiqu
AU2020454675A1 (en) 2020-06-24 2023-02-16 Sapreme Technologies B.V. Combination comprising an ADC or an AOC comprising a VHH, and a saponin or a ligand-saponin conjugate
WO2023121447A1 (fr) 2021-12-22 2023-06-29 Sapreme Technologies B.V. Conjugué d'un anticorps à domaine unique, d'une saponine et d'une molécule effectrice, composition pharmaceutique le comprenant, utilisation thérapeutique de ladite composition pharmaceutique

Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2001095935A1 (fr) * 2000-01-20 2001-12-20 Ottawa Health Research Institute Acides nucleiques immunostimulateurs permettant d'induire une reponse immunitaire th2
WO2002072631A2 (fr) * 2001-03-14 2002-09-19 Dakocytomation Denmark A/S Nouvelles constructions de molecules mhc, methodes d'utilisation de ces constructions a des fins de diagnostic et de therapie et utilisations de molecules mhc
US20050008717A1 (en) * 2002-12-27 2005-01-13 Einbond Linda Saxe Anti-neoplastic compositions comprising extracts of black cohosh

Family Cites Families (7)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US5191067A (en) * 1989-04-27 1993-03-02 The Salk Institute For Biological Studies Fibroblast growth factor conjugates
US5576288A (en) * 1989-04-27 1996-11-19 The Salk Institute For Biological Studies Fibroblast growth factor conjugates
WO1996005233A1 (fr) * 1994-08-11 1996-02-22 Shiseido Co., Ltd. Acide hyaluronique acetyle a faible poids moleculaire, emollient, et procedes d'obtention et de purification de cet acide
PT2266603E (pt) * 2000-10-18 2012-11-02 Glaxosmithkline Biolog Sa Vacinas tumorais
BR0311866A (pt) * 2002-06-11 2005-03-15 Panagin Pharmaceuticals Inc Composições de saponinas ou sapogeninas para terapia de câncer
US20050196409A1 (en) * 2003-09-24 2005-09-08 James Dao Compositions of botanical extracts for treating malignancy-associated changes
US20050175623A1 (en) * 2004-02-10 2005-08-11 Zheng-Pin Wang Saponins as anticancer agent

Patent Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2001095935A1 (fr) * 2000-01-20 2001-12-20 Ottawa Health Research Institute Acides nucleiques immunostimulateurs permettant d'induire une reponse immunitaire th2
WO2002072631A2 (fr) * 2001-03-14 2002-09-19 Dakocytomation Denmark A/S Nouvelles constructions de molecules mhc, methodes d'utilisation de ces constructions a des fins de diagnostic et de therapie et utilisations de molecules mhc
US20050008717A1 (en) * 2002-12-27 2005-01-13 Einbond Linda Saxe Anti-neoplastic compositions comprising extracts of black cohosh

Non-Patent Citations (3)

* Cited by examiner, † Cited by third party
Title
HEBESTREIT, P. ET AL: "Cytotoxic activity of the seeds from Agrostemma githago var. githago", PLANTA MED., vol. 69, 2003, pages 921 - 925, XP008042795 *
KELLER J ET AL: "Development of a novel molecular adapter for the optimization of immunotoxins", JOURNAL OF CONTROLLED RELEASE, ELSEVIER SCIENCE PUBLISHERS B.V. AMSTERDAM, NL, vol. 74, no. 1-3, 6 July 2001 (2001-07-06), pages 259 - 261, XP004297533, ISSN: 0168-3659 *
YESILADA E ET AL: "Effects of triterpene saponins from Astragalus species on in vitro cytokine release", JOURNAL OF ETHNOPHARMACOLOGY, ELSEVIER SCIENTIFIC PUBLISHERS LTD, IE, vol. 96, no. 1-2, 4 January 2005 (2005-01-04), pages 71 - 77, XP004657423, ISSN: 0378-8741 *

Cited By (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2007132481A2 (fr) * 2006-05-12 2007-11-22 Bharat Biotech International Limited Nouvelles molécules thrombolytiques et procédé associé
WO2007132481A3 (fr) * 2006-05-12 2008-06-26 Bharat Biotech Int Ltd Nouvelles molécules thrombolytiques et procédé associé
GB2451044A (en) * 2006-05-12 2009-01-14 Bharat Biotech Int Ltd Novel thrombolytic molecules and a process therefor
GB2451044B (en) * 2006-05-12 2010-06-16 Bharat Biotech Int Ltd Novel thrombolytic molecules and a process therefore
WO2021014026A2 (fr) 2019-07-25 2021-01-28 Sapreme Technologies B.V. Saponine bioactive liée à une fraction fonctionnelle

Also Published As

Publication number Publication date
JP2008505958A (ja) 2008-02-28
WO2006005581A3 (fr) 2006-04-27
CA2571739A1 (fr) 2006-01-19
WO2006005581A2 (fr) 2006-01-19
ES2395818T3 (es) 2013-02-15
US20080064762A1 (en) 2008-03-13
EP1623715B1 (fr) 2012-09-19

Similar Documents

Publication Publication Date Title
US20080064762A1 (en) Composition Comprising a Pharmacologically Active Agent Coupled to a Target Cell Specific Component, and a Saponin
Heisler et al. Combined application of saponin and chimeric toxins drastically enhances the targeted cytotoxicity on tumor cells
Lee et al. Macrophage activation by polysaccharide isolated from Astragalus membranaceus
Lee et al. Macrophage activation by polysaccharide fraction isolated from Salicornia herbacea
EP4241847A2 (fr) Molécule de liaison de ciblage cellulaire améliorée
Hemmerle et al. The antibody-based targeted delivery of TNF in combination with doxorubicin eradicates sarcomas in mice and confers protective immunity
Weng et al. The toxin component of targeted anti‐tumor toxins determines their efficacy increase by saponins
CN101507730B (zh) 表没食子儿茶素没食子酸酯与柔红霉素的组合物及其应用
Bachran et al. Epidermal growth factor receptor expression affects the efficacy of the combined application of saponin and a targeted toxin on human cervical carcinoma cells
EP1183050A2 (fr) Procede de fabrication d'un produit pharmaceutique injectable
AU2017389094B2 (en) Aptamer-drug conjugate and use thereof
Kim et al. Immunostimulating activity of crude polysaccharide extract isolated from Curcuma xanthorrhiza Roxb.
Citores et al. Targeting cancer cells with transferrin conjugates containing the non-toxic type 2 ribosome-inactivating proteins nigrin b or ebulin l
Bhargava et al. Targeted dianthin is a powerful toxin to treat pancreatic carcinoma when applied in combination with the glycosylated triterpene SO 1861
Ju et al. Licorice and its active compound glycyrrhizic acid ameliorates cisplatin-induced nephrotoxicity through inactivation of p53 by scavenging ROS and overexpression of p21 in human renal proximal tubular epithelial cells.
Wawrzynczak et al. Biochemical, cytotoxic and pharmacokinetic properties of an immunotoxin composed of a mouse monoclonal antibody Fib75 and the ribosome‐inactivating protein α‐sarcin from Aspergillus giganteus
Islam et al. Kaempferia rotunda tuberous rhizome lectin induces apoptosis and growth inhibition of colon cancer cells in vitro
Mohseni et al. Potent in vitro antitumor activity of B-subunit of Shiga toxin conjugated to the diphtheria toxin against breast cancer
Fang et al. Removing the sporoderm from the sporoderm-broken spores of Ganoderma lucidum improves the anticancer and immune-regulatory activity of the water-soluble polysaccharide
Venkatesan et al. Targeting HSP90/Survivin using a cell permeable structure based peptido-mimetic shepherdin in retinoblastoma
JP4914350B2 (ja) セラリシン（Ｓｅｒｒａｌｉｓｉｎｓ）のポリペプチド断片を含む医薬組成物
CN114222750A (zh) 改善治疗窗的皂苷衍生物
US20230277612A1 (en) Saponin derivatives for use in medicine
Davol et al. Saporin toxins directed to basic fibroblast growth factor receptors effectively target human ovarian teratocarcinoma in an animal model
Song et al. Enhanced anticancer effect of MAP30–S3 by cyclosproin A through endosomal escape

Legal Events

Date Code Title Description
PUAI Public reference made under article 153(3) epc to a published international application that has entered the european phase

Free format text: ORIGINAL CODE: 0009012

AK Designated contracting states

Kind code of ref document: A1

Designated state(s): AT BE BG CH CY CZ DE DK EE ES FI FR GB GR HU IE IT LI LU MC NL PL PT RO SE SI SK TR

AX Request for extension of the european patent

Extension state: AL HR LT LV MK

17P Request for examination filed

Effective date: 20060614

AKX Designation fees paid

Designated state(s): AT BE BG CH CY CZ DE DK EE ES FI FR GB GR HU IE IT LI LU MC NL PL PT RO SE SI SK TR

17Q First examination report despatched

Effective date: 20070112

REG Reference to a national code

Ref country code: DE

Ref legal event code: R079

Ref document number: 602004039344

Country of ref document: DE

Free format text: PREVIOUS MAIN CLASS: A61K0036360000

Ipc: A61K0047480000

GRAP Despatch of communication of intention to grant a patent

Free format text: ORIGINAL CODE: EPIDOSNIGR1

RIC1 Information provided on ipc code assigned before grant

Ipc: A61K 47/48 20060101AFI20120216BHEP

Ipc: A61P 35/00 20060101ALI20120216BHEP

Ipc: A61K 36/36 20060101ALI20120216BHEP

GRAS Grant fee paid

Free format text: ORIGINAL CODE: EPIDOSNIGR3

GRAA (expected) grant

Free format text: ORIGINAL CODE: 0009210

RIN1 Information on inventor provided before grant (corrected)

Inventor name: SUTHERLAND, MARK

Inventor name: MELZIG, MATTHIAS

Inventor name: HEISLER, IRING

Inventor name: FUCHS, HENDRIK

Inventor name: BACHRAN, CHRISTOPHER

Inventor name: HEBESTREIT, PHILIP

AK Designated contracting states

Kind code of ref document: B1

Designated state(s): AT BE BG CH CY CZ DE DK EE ES FI FR GB GR HU IE IT LI LU MC NL PL PT RO SE SI SK TR

REG Reference to a national code

Ref country code: GB

Ref legal event code: FG4D

REG Reference to a national code

Ref country code: CH

Ref legal event code: EP

REG Reference to a national code

Ref country code: IE

Ref legal event code: FG4D

REG Reference to a national code

Ref country code: AT

Ref legal event code: REF

Ref document number: 575658

Country of ref document: AT

Kind code of ref document: T

Effective date: 20121015

REG Reference to a national code

Ref country code: DE

Ref legal event code: R096

Ref document number: 602004039344

Country of ref document: DE

Effective date: 20121115

PG25 Lapsed in a contracting state [announced via postgrant information from national office to epo]

Ref country code: FI

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20120919

Ref country code: CY

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20120919

REG Reference to a national code

Ref country code: NL

Ref legal event code: VDEP

Effective date: 20120919

REG Reference to a national code

Ref country code: AT

Ref legal event code: MK05

Ref document number: 575658

Country of ref document: AT

Kind code of ref document: T

Effective date: 20120919

Ref country code: ES

Ref legal event code: FG2A

Ref document number: 2395818

Country of ref document: ES

Kind code of ref document: T3

Effective date: 20130215

PG25 Lapsed in a contracting state [announced via postgrant information from national office to epo]

Ref country code: SI

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20120919

Ref country code: SE

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20120919

Ref country code: GR

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20121220

PG25 Lapsed in a contracting state [announced via postgrant information from national office to epo]

Ref country code: NL

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20120919

Ref country code: EE

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20120919

Ref country code: BE

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20120919

Ref country code: CZ

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20120919

Ref country code: RO

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20120919

PG25 Lapsed in a contracting state [announced via postgrant information from national office to epo]

Ref country code: SK

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20120919

Ref country code: PT

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20130121

Ref country code: PL

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20120919

PG25 Lapsed in a contracting state [announced via postgrant information from national office to epo]

Ref country code: AT

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20120919

PLBE No opposition filed within time limit

Free format text: ORIGINAL CODE: 0009261

STAA Information on the status of an ep patent application or granted ep patent

Free format text: STATUS: NO OPPOSITION FILED WITHIN TIME LIMIT

PG25 Lapsed in a contracting state [announced via postgrant information from national office to epo]

Ref country code: BG

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20121219

Ref country code: DK

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20120919

26N No opposition filed

Effective date: 20130620

REG Reference to a national code

Ref country code: DE

Ref legal event code: R097

Ref document number: 602004039344

Country of ref document: DE

Effective date: 20130620

PG25 Lapsed in a contracting state [announced via postgrant information from national office to epo]

Ref country code: MC

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20120919

REG Reference to a national code

Ref country code: CH

Ref legal event code: PL

REG Reference to a national code

Ref country code: IE

Ref legal event code: MM4A

PG25 Lapsed in a contracting state [announced via postgrant information from national office to epo]

Ref country code: CH

Free format text: LAPSE BECAUSE OF NON-PAYMENT OF DUE FEES

Effective date: 20130731

Ref country code: LI

Free format text: LAPSE BECAUSE OF NON-PAYMENT OF DUE FEES

Effective date: 20130731

PG25 Lapsed in a contracting state [announced via postgrant information from national office to epo]

Ref country code: IE

Free format text: LAPSE BECAUSE OF NON-PAYMENT OF DUE FEES

Effective date: 20130713

PG25 Lapsed in a contracting state [announced via postgrant information from national office to epo]

Ref country code: TR

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT

Effective date: 20120919

PG25 Lapsed in a contracting state [announced via postgrant information from national office to epo]

Ref country code: HU

Free format text: LAPSE BECAUSE OF FAILURE TO SUBMIT A TRANSLATION OF THE DESCRIPTION OR TO PAY THE FEE WITHIN THE PRESCRIBED TIME-LIMIT; INVALID AB INITIO

Effective date: 20040713

Ref country code: LU

Free format text: LAPSE BECAUSE OF NON-PAYMENT OF DUE FEES

Effective date: 20130713

REG Reference to a national code

Ref country code: FR

Ref legal event code: PLFP

Year of fee payment: 12

REG Reference to a national code

Ref country code: FR

Ref legal event code: PLFP

Year of fee payment: 13

REG Reference to a national code

Ref country code: DE

Ref legal event code: R079

Ref document number: 602004039344

Country of ref document: DE

Free format text: PREVIOUS MAIN CLASS: A61K0047480000

Ipc: A61K0047500000

REG Reference to a national code

Ref country code: FR

Ref legal event code: PLFP

Year of fee payment: 14

REG Reference to a national code

Ref country code: FR

Ref legal event code: PLFP

Year of fee payment: 15

PGFP Annual fee paid to national office [announced via postgrant information from national office to epo]

Ref country code: IT

Payment date: 20220725

Year of fee payment: 19

Ref country code: GB

Payment date: 20220719

Year of fee payment: 19

Ref country code: ES

Payment date: 20220815

Year of fee payment: 19

Ref country code: DE

Payment date: 20220628

Year of fee payment: 19

PGFP Annual fee paid to national office [announced via postgrant information from national office to epo]

Ref country code: FR

Payment date: 20220727

Year of fee payment: 19

P01 Opt-out of the competence of the unified patent court (upc) registered

Effective date: 20230706

REG Reference to a national code

Ref country code: DE

Ref legal event code: R119

Ref document number: 602004039344

Country of ref document: DE

GBPC Gb: european patent ceased through non-payment of renewal fee

Effective date: 20230713

PG25 Lapsed in a contracting state [announced via postgrant information from national office to epo]

Ref country code: DE

Free format text: LAPSE BECAUSE OF NON-PAYMENT OF DUE FEES

Effective date: 20240201

Ref country code: GB

Free format text: LAPSE BECAUSE OF NON-PAYMENT OF DUE FEES

Effective date: 20230713

PG25 Lapsed in a contracting state [announced via postgrant information from national office to epo]

Ref country code: FR

Free format text: LAPSE BECAUSE OF NON-PAYMENT OF DUE FEES

Effective date: 20230731